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Diabetes primes neutrophils to undergo NETosis, which impairs wound healing

Nature Medicine volume 21, pages 815–819 (2015)Cite this article

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Abstract

Wound healing is impaired in diabetes, resulting in significant morbidity and mortality. Neutrophils are the main leukocytes involved in the early phase of healing. As part of their anti-microbial defense, neutrophils form extracellular traps (NETs) by releasing decondensed chromatin lined with cytotoxic proteins1. NETs, however, can also induce tissue damage. Here we show that neutrophils isolated from type 1 and type 2 diabetic humans and mice were primed to produce NETs (a process termed NETosis). Expression of peptidylarginine deiminase 4 (PAD4, encoded by Padi4 in mice), an enzyme important in chromatin decondensation, was elevated in neutrophils from individuals with diabetes. When subjected to excisional skin wounds, wild-type (WT) mice produced large quantities of NETs in wounds, but this was not observed in Padi4−/− mice. In diabetic mice, higher levels of citrullinated histone H3 (H3Cit, a NET marker) were found in their wounds than in normoglycemic mice and healing was delayed. Wound healing was accelerated in Padi4−/− mice as compared to WT mice, and it was not compromised by diabetes. DNase 1, which disrupts NETs, accelerated wound healing in diabetic and normoglycemic WT mice. Thus, NETs impair wound healing, particularly in diabetes, in which neutrophils are more susceptible to NETosis. Inhibiting NETosis or cleaving NETs may improve wound healing and reduce NET-driven chronic inflammation in diabetes.

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Figure 1: Diabetes or high glucose concentrations in vitro prime human and mouse neutrophils to undergo NETosis.
Figure 2: NETs are present in the wounds of WT mice.
Figure 3: PAD4 deficiency facilitates wound repair in normoglycemic mice.
Figure 4: PAD4 deficiency or DNase 1 treatment enhances wound healing in diabetic mice.

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Acknowledgements

We thank H. Ferris for advice on diabetes protocols; L. DeVita for selection of diabetic patients; P. Forbes (The Harvard Clinical and Translational Science Center, US National Institutes of Health (NIH) Award UL1 TR001102) for statistical advice; J.E. Cabral and S. Cifuni for valuable technical support; and L. Cowan for manuscript preparation assistance. This study was supported by the American Diabetes Association (Innovation Award 7-13-IN-44 to D.D.W.), the National Heart, Lung, and Blood Institute of the NIH (R01HL102101 to D.D.W.), the National Cancer Institute (R01CA136856 to Y.W.), the National Institute of Diabetes and Digestive and Kidney Diseases (R01DK031036 to C.R.K.) and a GlaxoSmithKline/Immune Disease Institute Alliance Fellowship (S.L.W.).

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Authors and Affiliations

  1. Program in Cellular and Molecular Medicine, Boston Children's Hospital, Boston, Massachusetts, USA

    Siu Ling Wong, Melanie Demers, Kimberly Martinod, Maureen Gallant & Denisa D Wagner

  2. Department of Pediatrics, Harvard Medical School, Boston, Massachusetts, USA

    Siu Ling Wong, Melanie Demers, Kimberly Martinod & Denisa D Wagner

  3. Department of Biochemistry and Molecular Biology, Center for Eukaryotic Gene Regulation, Pennsylvania State University, University Park, Pennsylvania, USA

    Yanming Wang

  4. Section of Clinical Research, Joslin Diabetes Center, Harvard Medical School, Boston, Massachusetts, USA

    Allison B Goldfine

  5. Section on Integrative Physiology and Metabolism, Joslin Diabetes Center, Harvard Medical School, Boston, Massachusetts, USA

    C Ronald Kahn

  6. Division of Hematology/Oncology,

    Denisa D Wagner

  7. , Boston Children's Hospital, Boston, Massachusetts, USA

    Denisa D Wagner

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Contributions

S.L.W. designed the study, performed the majority of the experiments, analyzed the data and wrote the manuscript; M.D. and K.M. performed experiments and analyzed data; M.G. provided expert technical assistance; Y.W. provided Padi4−/− mice and critical discussion of the work; A.B.G. provided clinical advice and selected diabetic patients for in vitro NETosis assays; C.R.K. provided helpful suggestions on experimental design and critical reading of the manuscript; D.D.W. designed the study, supervised the project and co-wrote the manuscript.

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Correspondence to Denisa D Wagner.

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Wong, S., Demers, M., Martinod, K. et al. Diabetes primes neutrophils to undergo NETosis, which impairs wound healing. Nat Med 21, 815–819 (2015). https://doi.org/10.1038/nm.3887

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